We are expanding the range of assay techniques that will allow us to understand the sulfation code in chondroitin sulfate glycosaminoglycan (CS-GAG) chains. These assay techniques take advantage of specific chromatography techniques (ion exchange, hydrophilicity) to separate the different disaccharides and monosaccharides that comprise the GAG chains. This is the only technique capable of doing this. We have also initiated mass specrtrophotometric analysis of these separated GAG chains to begin to determine the sequence of sulfations on the different parts of the GAG chain. We have developed assays of growth cone response to chondroitin sulfate proteoglycans. We have completed studies that determined that the activity of myosin II molecular motors are essential for this growth cone turning. Both myosin IIA and IIB appear to be involved in growth cone turning. Modification of their activity by drugs may ultimately prove to be a useful therapeutic approach for treatment of brain injury. Because many of these mechanisms are also found in injury to heart and blood vessels, these approaches may have a more general applicability. We have instituted studies to develop a systems biology approach to CSPG actions. We have identified a number of proteins whose phosphorylation is changed rapidly as neurons are exposed to CSPGs. We have confirmed the changes in several of these with Western blotting techniques. We are now developing an approach to validate changes in the several different signal transduction pathways. We have inititated collaborative studies that have identified the Nogo receptor as a new receptor for CSPGs. In addition our data point to an additional receptor that binds bioactive CSPGs.